A total of 21 aquatic plant species were collected from Bantimurung waterfall areas, South Sulawesi, Indonesia, in October 2017. These plant materials were subjected to both species and phytochemicals identification. The aims of this study were to determine the species or taxonomic rank of Indonesian aquatic plants collected from Bantimurung waterfall, South Sulawesi, Indonesia and to identify their chemical compounds (phytochemicals) as a candidate for new herbal medicine. Plant genetic materials used in this study were collected from Bantimurung Bulusaraung waterfall and were then identified based on standard botanical techniques for species identification in the Herbarium Bogoriense, Research center for Biology Indonesian Institute of Science (LIPI), Cibinong, West Java. The samples were subjected to the phytochemistry screening such as alkaloids, flavonoids, tannins, saponins, glycosides, terpenoids and anthraquinone followed the procedures of Indonesian Materia Medika and Harborne.  Results showed that all collected aquatic plant samples were able to be identified, including their species names. Phytochemical screening of each sample revealed the presence of glycoside in all of the tested species. However, no alkaloids, anthraquinones, and terpenoids were observed in those tested plant samples. Of the total 21 aquatic plants, 14 species contained flavonoids, 8 species contained phenol compound, and 10 species contained saponins. Among these species Donnax canniformis possessed good antioxidant activity, which correlated to its total phenolic and flavonoid contents. Our results would be beneficial for any future effort in the development of new herbal drugs derived from aquatic plants.

Promoter regulates expression level of foreign gene in transgenic organism. This study was performed to select a suitable promoter as the fi rst step towards production of valuable trait-enhanced seaweed by transgenic technology. Green fl uorescent protein (GFP) gene was used as a reporter to determine the activity of promoter in seaweed Kappaphycus alvarezii. GFP gene constructs driven by cytomegalovirus (pCMV-GFP), caulifl ower mosaic virus (pCaMV-GFP), medaka β-actin (pmBA-GFP) and Japanese fl ounder keratin (pJfKer-GFP) promoters were introduced by electroporation method. Electroporation was performed using a gene pulser (BIORAD) with voltage of 300 V, pulse length of 0.5 ms, pulse numbers of 4, and pulse interval of 0.1 s. Promoter activity was determined by analyzing GFP gene expression level using a fl uorescent microscope. The results showed that CMV regulated highest number of fi lament callus (34.10%±1.49) expressing GFP at medium to strong fl uorescence levels. CaMV promoter had relatively similar activity with CMV, but lower number of fi lament callus expressing GFP (10.48%±0.25). mBA promoter drove GFP expression at medium level and similar number of fi lament callus (8.85%±2.31) expressing GFP with CaMV, while JfKer promoter had lowest activity by means in number of fi lament callus expressing GFP (4.79%±0.26) and GFP expression level. PCR analysis for transgenic confi rmation showed a DNA band of PCR product from pCMV-GFP and pCaMV-GFP expressing fi lament callus in the same size (about 0.6 kb) with positive control of plasmid. Thus, CMV and CaMV promoters was an appropriate promoter and foreign gene could be transferred to fi lament callus by electroporation method. Combining this achievement with developing a culture method of fi lament callus to be thallus, stable transgenic breeding in K. alvarezii can be feasible.

Untuk mendukung program transgenesis pada rumput laut, embrio somatik dapat digunakan sebagai material untuk transfer gen baik secara individu sel ataupun kluster sel embriogenik, sehingga mempercepat keberhasilan dengan peluang transformasi yang lebih tinggi. Penelitian ini bertujuan untuk mengkaji induksi kalus rumput laut K. alvarezii untuk produksi sel embrio somatik (e.s.) dengan beberapa rasio zat pengatur tumbuh (ZPT) dan konsentrasi agar media induksi, sampai sel menjadi filamen. Penelitian terdiri atas dua tahap: Tahap (1) induksi kalus, dengan rasio ZPT asam indol asetat (IAA):kinetin = 0,5:0,0 mg/L; 1,0:1,0 mg/L; dan 2,0:0,2 mg/L dengan konsentrasi agar media induksi = 0,6%; 0,8%; 1,0%; dan 1,5%. Tahap (2) regenerasi massa sel e.s., dengan rasio IAA:kinetin = 0,1:1,0 mg/L; 0,0:0,1 mg/L dan tanpa ZPT dengan konsentrasi agar media = 0,4%; 0,6%; dan 0,8%. Untuk perkembangan sel-sel e.s. lebih lanjut dipelihara pada kultur cair. Hasil penelitian menunjukkan pada tahap induksi kalus, rasio IAA: kinetin = 1:1 mg/L dengan konsentrasi agar media 0,8% dan 1,0% menghasilkan persentase induksi kalus tertinggi (90%). Pada tahap regenerasi massa sel e.s., ZPT tidak berpengaruh terhadap perkembangan massa sel e.s., di mana tanpa ZPT dengan konsentrasi agar 0,6% memperlihatkan perkembangan tertinggi (rata-rata diameter massa sel 5 mm). Pada media cair, perkembangan sel e.s. dari single cell ukuran 3-4 mm menjadi filamen-filamen ukuran rata-rata 0,5 mm dapat dicapai dalam satu bulan kultur. Keberhasilan produksi sel e.s. K. alvarezii, selain sebagai material untuk transfer gen juga dapat dijadikan acuan dalam produksi benih rumput laut kultur jaringan.To support the program of seaweed transgenesis, somatic embryo can be used as a materials for gene transfer purpose either by individual or cluster of cells in accelerating the higher rate of transformation. This research aims to study the callus induction of seaweed K. alvarezii for production of somatic embrio (s.e) cell by different ratio of growth regulators (GR) and agar media concentrations. The study consists of two stages: stage (1) callus induction to the cells filament using GR of indol acetic acid (IAA):kinetin in ratio of  0.5:0.0 mg/L; 1.0:1.0 mg/L; and 2.0:0.2 mg/L on the media agar concentration of 0.6%; 0.8%; 1.0%; and 1.5%, and stage (2) regeneration of the cell mass of s.e, using GR of IAA:kinetin in ratio of 0.1:1.0 mg/L; 0.0:0.1 mg/L on the media agar concentration of 0.4%; 0.6%; and 0.8%. For further maintenance, the s.e cells were cultured in liquid media. The results of callus induction showed that the ratio of IAA:kinetin (1:1) on the media agar concentration of 0.8% and 1.0% produced the highest callus induction (90%). The study of mass cell regeneration of s.e showed did not different of GR IAA:kinetin ratio of the cell mass development, but the media agar concentration of 0.6% and 0.4% showed the higher growth of cell mass (in diameter size of 4-5 mm). The development of cells culture of s.e from the size of 3-4 mm to filament of 0.5 mm could be reached in one month of culture period. The success of K. alvarezii s.e production will be helpful not only as a material for gene transfer but also as a reference on tissue culture for seed production of seaweed.

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Increasing of kappa (κ)-carrageenan content in Kappaphycus alvarezii seaweed is potentially be achieved by applying transgenesis technology. This study was performed to obtain a construction of  κ-Carrageenase gene and Agrobacterium tumefaciens to carry those construction genes.  The κ-Carrageenase (κ-Car) gene was involved in κ-carrageenan biosynthesis. The κ-Car gene sequence was ligated between the 35S CaMV promoter and tNos terminator sequences to generate pMSH/κ-Car expression vector. Transformation of pMSH/κ-Car plasmid to Escherichia coli was performed by heat-shock method, and to Agrobacterium tumefaciens by tri-parental mating method. The results showed that several colonies of E. coli and A. tumefaciens grew in the selective culture mediums containing antibiotic. PCR analysis using primers 35S-Forward and tNos-Reverse with DNA template from those bacterial colonies resulted DNA fragment of about 2,000 bp, the same as the total length of 35S CaMV promoter, κ-Car gene and tNos terminator sequences. Therefore, the construction of pMSH/κ-Car gene was succeeded and a colony of A. tumefaciens transformant carrying pMSH/κ-Car plasmid was successfully produced.                                                                                   Keywords:  Agrobacterium tumefaciens, kappa(κ)-Carrageenase gene, transgenesis, vector